Combinatorial approach to develop novel pre-therapeutic agents targeting virulence factors essential to clinically relevant pathogens

开发针对临床相关病原体必需毒力因子的新型治疗前药物的组合方法

• 批准号: 10681469
• 负责人: Hanh Ngoc Lam
• 金额: $ 24.07万
• 依托单位: VIRGINIA POLYTECHNIC INST AND ST UNIV
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-09 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10681469
• 关键词: Active Sites; Affinity; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Aspartate; Bacteria; Binding; Binding Proteins; Biochemical; Biological; Biological Assay; Biology; C-terminal; Calorimetry; Cause of Death; Cell Death; Cell membrane; Cells; Cessation of life; Chemicals; Collaborations; Communicable Diseases; Computer Assisted; Consumption; Data; Development; Drug resistance; ESKAPE pathogens; Enzyme Activation; Fluorescent Probes; Foundations; Future; Goals; Gram-Negative Bacteria; Health; Health Care Costs; Host Defense Mechanism; Hybrids; Infection; Inflammation; Knowledge; Learning; Legal patent; Libraries; Medicine; Membrane; Mentors; Methods; Microbe; Molecular Target; Mus; N-substituted Glycines; Natural Products; Nosocomial Infections; Outcome; Pathogenesis; Pathogenicity; Pathology; Peptides; Peptoids; Periodicity; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phase; Phenotype; Phospholipase A2; Process; Productivity; Proteins; Pseudomembranous Colitis; Pseudomonas; Pseudomonas aeruginosa; Regulation; Research; Resistance; Resolution; Series; Serine; Serine Hydrolase; Specificity; Structure; Structure-Activity Relationship; Techniques; Therapeutic; Time; Titrations; Type III Secretion System Pathway; United States; Virulence; Virulence Factors; Virulent; Work; X-Ray Crystallography; acute infection; algorithm development; antibiotic resistant infections; antimicrobial; antimicrobial drug; appendage; clinically relevant; cofactor; combat; combinatorial; commensal microbes; cost; cost effective; design; drug candidate; drug discovery; extracellular; genetic approach; global health; healthcare-associated infections; high throughput screening; inhibitor; innovation; insight; lead optimization; macromolecule; marine; microbial; microbiome; microbiota; multidrug-resistant Pseudomonas aeruginosa; mutant; novel; novel strategies; novel therapeutics; pathogen; pathogenic bacteria; pressure; prevent; priority pathogen; screening; skills; small molecule; targeted treatment; therapeutic target; trait; virtual screening

Project Summary/Abstract Antibiotic resistance is a serious global health threat. Current antibiotics target essential bacterial processes and impose strong selective pressure for resistance. Upon antibiotic treatment, the healthy microbiota are reduced in both number and diversity, leading to serious health consequences such as Clostridium difficile colitis. Moreover, resistant traits can be transferred to other microbes, leading to the spread of antibiotic resistance. Using virulence blockers to target specific pathogenicity mechanisms, while leaving the microbiota intact, is a promising strategy to reduce resistance. This proposal will identify molecular target of a newly discovered the type III secretion system (T3SS) inhibitor, and explore their modes of action for further optimization and development. I will assess structure – activity relationship to optimize the T3SS inhibitors, cyclic pepeptomers, and use affinity based method to identify their molecular targets in Pseudomonas aeruginosa. Besides, target-based drug discovery offers the advantage of being low cost and less time consuming. With the availability of high-resolution structure and development of algorithms to predict binding affinity and poses of small molecules to its protein target, virtual screening can provide lead for optimization. ExoU, an effector with phospholipase A2 activity, is the major effector in P. aeruginosa, one of six ESKAPE pathogens which cause the majority of nosocomial infections in the U.S. and “escape” antimicrobial drugs. ExoU has a serine/aspartate catalytic dyad and a separate cofactor-binding domain required for activation of the enzyme. ExoU is highly toxic, associated with acute infection, antibiotic resistance and severe outcome in patents. Delay ExoU expression can increase mice survival. Thus we set out to find ExoU inhibitors as a strategy to treat acute infection and reduce resistance. We will identify ExoU inhibitors that 1) inhibit enzymatic activity by targeting its catalytic residue serine, or 2) bind to the membrane localization domain which will prevent ExoU's activation by virtual screening. The inhibitors that show binding affinity to ExoU in isothermal titration calorimetry assay, and prevent cell death caused by ExoU will be chosen for optimization. Structure of inhibitor-bound proteins will be solved using X-ray crystallography. I believe that my team of mentors (Drs. Stone and Ottemann), advisors (Dr. Rubin, expert in X-ray crystallography; Dr. Jacobson, expert in computer- aided drug discovery) and collaborators (Dr. Lokey, an expert in macromolecule synthesis, Drs. Crews and Linington, natural product chemists) will provide me support to successfully carry out this project. With the biochemical techniques I will learn, the structures and new inhibitors I will obtain in the K99 phase, I will then collaborate with Dr. Shaw (medicinal chemist) and Dr. Jacobson to optimize the candidate hits in my independent phase. This project extends my skill set in biochemical methods and has the potential to provide substantial momentum towards drug discovery, and development. These will serve as the foundation of a R01 proposal to be prepared upon the completion of the main stages of this research plan.

项目总结/文摘